Phytotoxicity of Alkaloids, Coumarins and Flavonoids Isolated from 11

Home , Meliaceae, Rutaceae

Phytochemistry Letters 8 (2014) 226–232

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Phytochemistry Letters

jo urnal homepage: www.elsevier.com/locate/phytol

Phytotoxicity of alkaloids, coumarins and ﬂavonoids isolated from

11 species belonging to the Rutaceae and Meliaceae families

a b b a

Liliane Nebo , Rosa M. Varela , Jose´ M.G. Molinillo , Olı´via M. Sampaio ,

a a a

Vanessa G.P. Severino , Cristiane M. Cazal , Maria Fa´tima das Grac¸as Fernandes ,

a b,

Joa˜o B. Fernandes , Francisco A. Macı´as *

Laboratory of Natural Products, Department of Organic Chemistry, Federal University of Sa˜o Carlos, 13560-970 Sa˜o Carlos (SP), Brazil

Allelopathy Group, Department of Organic Chemistry, Institute of biomolecules (INBIO), School of Sciences University of Ca´diz, 11510 Puerto Real (Ca´diz),

Spain

A R T I C L E I N F O A B S T R A C T

Article history: Meliaceae and Rutaceae families are known for the high diversity of their secondary metabolites, which

Received 7 February 2014

include many groups that represent a rich source of structural diversity, and are good candidates as

Received in revised form 14 February 2014

sources of allelochemicals that could be useful in agriculture. In the work described here the bioactivity

Accepted 21 February 2014

proﬁles were evaluated for 3 alkaloids (1–3), 12 coumarins (4–15), 2 phenylpropanoic acid derivatives

Available online 11 March 2014

(16 and 17) and 14 ﬂavonoids (18–31) from 11 species belonging to the Meliaceae and Rutaceae families.

All compounds were tested in the wheat coleoptile bioassay and those that showed the highest activities

Keywords:

were tested on the STS (Standard Target Species) Lepidium sativum (cress), Lactuca sativa (lettuce),

Allelopathy

Lycopersicon esculentum (tomato), and Allium cepa (onion).

Phytotoxicity

Meliaceae Most of the isolated compounds showed phytotoxic activity and graveoline (3), psoralen (8), and

Rutaceae ﬂavone (18) were the most active, with bioactivity levels similar to that of the commercial herbicide

Alkaloids Logran . The results indicate that these compounds could be involved as semiochemicals in the

Coumarins allelopathic interactions of these plant species.

1. Introduction 2011), and insecticidal (Bermu´ dez-Torres et al., 2009; Li et al.,

2009) properties amongst others.

Meliaceae and Rutaceae families are known for their high Coumarins are a large group of compounds that are widely

diversity of secondary metabolites, including many groups that distributed in many plant species and they have a wide spectrum of

represent a rich source of structural diversity. Alkaloids, coumar- biological activities, including insecticidal (Pavela and Vrchotova´,

ins, ﬂavonoids, terpenes and limonoids are the largest groups of 2013), anticancer (Harada et al., 2010), Alzheimer’s disease (Anand

secondary plant metabolites and they show remarkable biological et al., 2012) and antimicrobial (Al-Amiery et al., 2012; Guan et al.,

activities and have potential medicinal value (Tan and Luo, 2011; 2011; Souza et al., 2005; Godoy et al., 2005; Sardari et al., 1999).

Champagne et al., 1992; Da Silva et al., 1984; Da Silva and Gottlieb, Flavonoids are also an important group of compounds and are

1987). widely found in the plant kingdom. They have a diverse range of

The biological activity of the alkaloids has been actively signiﬁcant bioactivities, including antioxidant (Wolfe and Liu,

explored and has been found to span a variety of properties 2008), allelopathic (Treutter, 2006), gastroprotective (Mota et al.,

(Michael, 2007, 2005). These include anticancer (Duarte et al., 2009), anticancer (Pick et al., 2011; Benavente-Garcı´a and Castillo,

2010; Zhai et al., 2012), antiparasitic (Silva et al., 2012; Mbeunkui 2008; Marchand, 2002), anti-inﬂammatory (Garcı´a-Lafuente et al.,

et al., 2012; Mishra et al., 2009; Santos et al., 2009), anti- 2009; Kim et al., 2004), and antimicrobial (Cushnie and Lamb,

inﬂammatory (Souto et al., 2011), antimicrobial (Joosten and Veen, 2011; Salas et al., 2011) along with other effects.

The increased interest in crop protection, along with the

extensive use of agrochemicals and the problems associated with

their use, has led to the search for new biologically active natural

products (Dayan et al., 2009). The discovery of new allelochemicals

* Corresponding author. Tel.: +34 956 01 2770; fax: +34 956 016193.

is an attractive alternative to current conventional herbicides used

E-mail address: [email protected] (F.A. Macı´as).

http://dx.doi.org/10.1016/j.phytol.2014.02.010

L. Nebo et al. / Phytochemistry Letters 8 (2014) 226–232 227

OCH3 O

O O O O O O

O O H3CO N N O O O

CH3 O O

1 R=H 3 4 O 5 6

2 R = OCH3

R O O O O HO O O O O O O O O

7 8 R=H R'= H 11 12

9 R =OCH3 R'= H

10 R = H R'= OCH3 OH OCH3 OCH3 O HO O O HO R

HO OH

HO O O O O O O OCH

HO O O 3

13 14 15 16 R = OH

17 R =OCH3

Fig. 1. Structures of the alkaloids (1–3) and coumarins (4–17) isolated from Meliaceae and Rutaceae families.

for weed control. Commercial herbicides have caused changes in 2 and 9 from roots and aerial parts of Ruta graveolens (Paulini et al.,

populations of invasive species and environmental pollution and 1989; Masuda et al., 1998); 4 from roots of Citrus sinensis grafted

they have also enhanced the resistance phenomenon (Macı´as et al., on Citrus limonia (Cazal et al., 2009); 5 from stems and leaves of

2007). In this context, Meliaceae and Rutaceae are good candidates Raunia resinosa (Velozo et al., 1997); 7 from the fruit of Swinglea

to provide a source of allelochemicals for future use in agriculture. glutinosa (Santos, 2005); 15, 16 and 17 from stem and taproots of

The aim of the work described here was to evaluate the Hortia oreadica (Braga et al., 2012); 18, 23 and 24 from the fruit,

bioactivity proﬁles of three alkaloids (1–3), twelve coumarins (4– branches and leaves of C. fruticosa Bl. (Leite et al., 2009); 21 from

15), two phenylpropanoic acid derivatives (16 and 17) (Fig. 1) and the fruit of Neoraputia magniﬁca (Tomazela et al., 2000; Passador

fourteen ﬂavonoids (18–31) (Fig. 2) from eleven species belonging et al., 1997); 22 from leaves of Neoraputia alba (Arruda et al.,

to the Meliaceae and Rutaceae families. 1993); 25 and 29 from stems and leaves of Neoraputia paraensis

(Moraes et al., 2003); 26 and 27 from peel; and 31 from the fruit of

2. Results and discussion Murraya paniculata (Ferracin et al., 1998). Metabolites 13, 14, 19,

20, 28 and 30 were obtained from commercial sources in order to

2.1. Isolation of compounds carry out bioassays.

Compounds 1, 3, 8, 11 and 12 were isolated from the roots and

The extraction, isolation and identiﬁcation (NMR, MS, IR, UV aerial parts of R. graveolens, and 10 and 15 were obtained from

data) of the following compounds has been described previously: taproots of H. oreadica as described in Section 3.

Fig. 2. Structures of the ﬂavonoids (18–31) isolated from Meliaceae and Rutaceae families.

228 L. Nebo et al. / Phytochemistry Letters 8 (2014) 226–232

Fig. 3. Bioactivities obtained in the etiolated wheat coleoptile bioassay for compounds 1–17. Values are expressed as percentage differences from the control and are not

a b

significantly different with P > 0.05 for the Mann–Whitney test. Values significantly different with P < 0.01. Values significantly different with 0.01 < P <0.05.

2.2. Coleoptile bioassay results 10 and 15) leads to better results. However, the presence of prenyl

groups bonded to C-3 does not lead to a decrease in the activity (8

The etiolated wheat coleoptiles bioassay was used as an initial vs. 11 and 12). Simple coumarins (13 and 14) and phenylpropanoic

approach to evaluate the phytotoxicity of these compounds since it acid derivatives (16 and 17) showed inhibition effects of between

is a rapid test (24 h) that is sensitive to a wide range of bioactive 70% and 100% at the highest concentration, although the

substances (Cutler et al., 2000), including plant growth regulators, activities of these compounds decrease rapidly with dilution.

herbicides (Cutler, 1984), antimicrobials, mycotoxins and assorted Regarding ﬂavonoids, the most active compounds were ﬂavone

0 0

pharmaceuticals (Jacyno and Cutler, 1993). The results are shown (18) and 3 ,4 -methylenedioxy-5,7-dimethoxyﬂavone (21), which

in Figs. 3 and 4, where negative values signify inhibition, positive gave values of around 98% and 76% at 1 mM, respectively.

0 0 0

values denote activation and zero represents control. 3 ,4 ,5,5 ,7-Pentamethoxyﬂavone (23) did not show the highest

All alkaloids assayed were active. Evolitrine (1) and graveoline levels of inhibition at 1 mM and 300 mM (47% and 57%,

(3) presented the most consistent proﬁles, with levels of activity respectively), but the bioactivity was maintained upon dilution

higher than – 85% at the ﬁrst three concentrations tested (1 mM, (51%, 100 mM; 44%, 30 mM and 44%, 10 mM) (Fig. 4). There is

300 mM and 100 mM) (Fig. 3). Kokusagine (2) differs from 1 in the no clear substitution pattern that would indicate whether

presence of an additional methoxyl group at C-6 but it only compounds are active or not. The introduction of hydroxyl and

showed relevant activity levels at 1 mM and 300 mM (–94% methoxyl groups seems to inﬂuence the bioactivity and small

and –70%). structural differences can change the activity markedly, as can be

Regarding coumarins, xanthyletin (4) was the most active of the observed for 21 and 22 (Macı´as et al., 1997).

pyranocoumarins (4–7, 15). This compound completely inhibited

coleoptile elongation at 1 mM, 300 mM and 100 mM and the 2.3. Phytotoxicity bioassay

bioactivity was retained upon dilution (–68%, 30 mM; 10 mM). Of

the furanocoumarins, psoralen (8), chalepin (11) and chalepensin The most active compounds, alkaloids 1 and 3, pyranocoumar-

(12) showed good levels of activity. The most active compound was ins 4 and 8, furanocoumarins 11 and 12 and the ﬂavones 18, 21 and

12, which completely inhibited the coleoptile elongation at 1 mM 23 were selected for phytotoxicity evaluation on the standard

and 300 mM with the bioactivity maintained upon dilution (76%, target species (STS) Lepidium sativum (cress), Lactuca sativa

100 mM; 72%, 30 mM and 46%, 10 mM). These results allow (lettuce), Lycopersicon esculentum (tomato), and Allium cepa

some structural correlations to be made. For example, linear (onion). The commercial herbicide Logran was used as internal

pyranocoumarins and furanocoumarins are preferred to angular standard (Macı´as et al., 2000). The results of the bioassay are

compounds (4, 8, 11 and 12 vs. 5, 7 and 15). Furthermore, the presented in Figs. 5 and 6, where data are presented as percentage

absence of alkyl groups at C-8 (4 vs.6) and methoxyl groups (8 vs. 9, differences from the control. The concentrations tested were

Fig. 4. Bioactivities obtained in the etiolated wheat coleoptile bioassay for compounds 18–31. Values are expressed as percentage differences from the control and are not

a b

significantly different with P > 0.05 for the Mann–Whitney test. Values significantly different with P < 0.01. Values significantly different with 0.01 < P <0.05.

L. Nebo et al. / Phytochemistry Letters 8 (2014) 226–232 229

Fig. 5. Effects of the commercial herbicide Logran and 1, 3, 4, 8, 11 and 12 on growth of standard target species. Values are expressed as percentage differences from the

a b

control and are not significantly different with P > 0.05 for the Mann–Whitney test. Values significantly different with P < 0.01. Values significantly different with

0.01 < P <0.05.

identical to those in the coleoptile bioassay, with the exception of active and they inhibited root growth at all concentrations with

1, for which the highest concentration was 300 mM. similar levels to the herbicide Logran (positive control).

The least affected parameter was germination, apart from Graveoline (3) and psoralen (8) showed the best results on

compounds 3 and 8, which inhibited germination of L. sativum, L. shoot growth, with values of 75% and 69%, respectively, at

esculentum and A. cepa. the highest concentration (10 M) (Fig. 5). Compounds 3 and

Regarding the dicotyledonous species, L. sativum, the 8 also affected the germination (62% and 83%, respectively,

compounds evolitrine (1) and chalepensin (12) were the most at 1 mM).

230 L. Nebo et al. / Phytochemistry Letters 8 (2014) 226–232

Fig. 6. Effects of the commercial herbicide Logran and 18, 21 and 23 on growth of L. sativum. Values are expressed as percentage differences from the control and are not

a b

significantly different with P > 0.05 for the Mann–Whitney test. Values significantly different with P < 0.01. Values significantly different with 0.01 < P <0.05.

Compound 21 was the most active ﬂavone on germination and was collected in Sa˜o Carlos, SP, Brazil; Cipadessa fruticosa Bl and N.

growth of L. sativum and it showed similar levels of phytotoxicity magniﬁca were collected in Vic¸osa, MG, Brazil; N. alba and R.

to the herbicide Logran (Fig. 6). The effect on other species was resinosa (Nees et. Mart.) were collected in Cachoeiro do Itapemirim,

not significant. The behavior of 18 and 23 was not significant on Espı´rito Santo State, Brazil. The plants were identified by Dr. Jose´ R.

STS species. Pirani from the Department of Botany, University of Sa˜o Paulo and

Regarding L. sativa, the most active compounds were 8 and 12 vouchers were deposited at the Herbarium at the same Depart-

and these affected the root growth at the highest concentration ment.

(65% and 56%, respectively, at 1 mM). The growth of L. S. glutinosa (Bl.) Merr. was collected in Campinas, SP, Brazil, and

esculentum and A. cepa was also affected by the tested compounds identiﬁed by Prof. Dr. Maria Ineˆs Salgado. A voucher specimen is

– especially in the cases of 3 and 8, which inhibited both deposited at the Herbarium of the Botany Department, Federal

parameters in both species. University of Sa˜o Carlos (UFSCar) as number 7110.

Hale et al., 2004 demonstrated that 3 affected the growth of the

aquatic plant Lemma paucicostata at 100 mM and caused tissue 3.2. Extraction and isolation

degradation at 250 mM and above. On the other hand, compounds

4, 9, and 10 have been previously tested on L. sativa and it was 3.2.1. Isolation of 1, 3, 8, 11 and 12 from Ruta graveolens

proposed that these compounds are responsible for the allelo- Dried roots (1.6 kg) and aerial parts (3.4 kg) were extracted

pathic activity of Pilocarpus goudotianus (Macı´as et al., 1993). with cold ethanol using a homogenizer for 3 days. The ethanolic

Although xanthyletin (5) and chalepin (11) have shown only solution was concentrated under reduced pressure and the crude

moderate phytotoxicity levels on STS species, they inhibited the extract of aerial parts was chromatographed using silica gel (CC) to

root growth of Amaranthus hypochondriacus (Anaya et al., 2005). afford the following fractions: hexane (13.8 g), CH2Cl2 (25.0 g) and

Sampaio et al., 2012 evaluated the photosynthesis inhibition MeOH (95.0 g). The liquid/liquid partition technique was used to

potential of furanocoumarins and highlighted chalepin (11) as a fractionate the root extract into the following fractions: hexane

photosynthesis inhibitor. (1.53 g), CH2Cl2 (1.55 g), EtOAc (1.87 g) and water (1.99 g). The

We propose Meliaceae and Rutaceae to be good candidates to CH2Cl2 fraction from the aerial parts was chromatographed on SiO2

provide sources of allelochemicals that may be useful in (70–230 mesh; 5.3 cm 21.0 cm) in vacuum and eluted with

agriculture. The isolated compounds showed phytotoxic activity hexane (A), CH2Cl2 (B) and MeOH (C) to yield three fractions. The

and graveoline (3), psoralen (8) and ﬂavone (18) were the most CH2Cl2 fraction (B) from the aerial parts was fractionated on SiO2,

active, with bioactivity levels similar or even better than the eluted with hexane, CH2Cl2, Me2CO and MeOH to afford 20

commercial herbicide Logran . The bioactivities of these com- fractions from which 8 (110.0 mg) and 12 (230.0 mg) were

pounds indicate that these products could also be involved as isolated. These new fractions were further puriﬁed by HPLC

semiochemicals in the allelopathic interactions of these plant [Phenyl-Hexyl (0.7 30 cm) and particle size of 10 mm], with

species. elution in the isocratic mode: MeOH:CH2Cl2, 1:1 (v/v), ﬂow

rate = 3.0 mL min . Detection (Shimadzu SCL-10A) was carried

3. Experimental out at l = 254 and 365 nm and compound 11 (52.2 mg) was

isolated. The MeOH fraction (C) from the aerial parts was puriﬁed

3.1. Plant material on Sephadex LH-20 in isocratic mode (MeOH:CH2Cl2, 1:1 (v/v)) and

compound 3 (35.2 mg) was isolated. The CH2Cl2 fraction from the

R. graveolens was collected in Akai Ranch, located in Atibaia, Sa˜o roots was fractionated on SiO2, eluted with hexane, CH2Cl2, Me2CO

Paulo State, Brazil; H. oreadica Groppo was collected in Forest and MeOH. A total of 30 fractions were collected and 1 (13.2 mg)

Reserve Adolpho Ducke, Itacoatiara, Amazonas State, Brazil; C. was isolated.

sinensis grafted on C. limonia was collected in Estac¸a˜o Experimental The structures of the compounds were established by

de Citricultura do Instituto Agronoˆmico de Campinas, Cordeir- comparison of their spectroscopic and physical data with those

o´ polis, SP, Brazil; R. resinosa (Nees et. Mart.) was collected in reported in the literature (Hongwei et al., 2010; Masuda et al.,

Cachoeiro do Itapemirim, Espı´rito Santo State, Brazil; M. paniculata 1998; Ngadjui et al., 1998; Oliveira et al., 1996; Kumar et al., 1995).

L. Nebo et al. / Phytochemistry Letters 8 (2014) 226–232 231

3.2.2. Isolation of 10 and 15 from H. oreadica Petri dish. Four replicates were used for tomato, cress, onion and

The taproots (3.3 kg) of H. oreadica were dried carefully by lettuce (80 seeds). After adding seeds and aqueous solutions, Petri

forced air at 40 8C and the sample was powdered. The sample was dishes were sealed with Paraﬁlm to ensure closed-system models.

extracted to give the crude extracts as follows: hexane (86.0 g), Seeds were further incubated at 25 8C in a Memmert ICE 700

CH2Cl2 (68.0 g) and MeOH (244.0 g). The CH2Cl2 extract was controlled environment growth chamber in the dark. Bioassays

chromatographed on SiO2 under vacuum, eluted with hexane took 4 days for cress, 5 days for lettuce and tomato and 7 days for

(1.0 L), CH2Cl2 (1.0 L), EtOAc (3.0 L) and MeOH (1.0 L) to yield 4 onion.

fractions. The EtOAc fraction was chromatographed on SiO2 (230– After growth, plants were frozen at 10 8C for 24 h to avoid

400 mesh; 4.0 cm 24.0 cm), eluted with a hexane/MeOH subsequent growth during the measurement process. The com-

gradient to give 20 new fractions. Fraction 8 was puriﬁed on mercial herbicide Logran , a combination of 2-tert-butylamino-4-

SiO2 (230–400 mesh; 5.0 cm 30.0 cm) eluted with hexane/EtOAc ethylamino-6-methylthio-1,3,5-triazine (terbutryn, 59.4%) and 1-

with increasing polarity to afford compound 5 (128.0 mg). Fraction [2-(2-chloroethoxy)phenylsulfonyl]-3-(4-methoxy-6-methyl-

13 was chromatographed on SiO2 (230–400 mesh; 1,3,5-triazin-2-yl)urea (triasulfuron, 0.6%), was used as an internal

4.0 cm 30.0 cm) using the same methodology described above reference according to a comparison study reported previously

to yield compound 10 (4.0 mg). (Macı´as et al., 2000).

Structures of compounds were established by comparison of The herbicide was used at the same concentrations (1 mM, 300,

their spectroscopic and physical data with those reported in the 100, 30, 10 mM) and under the same conditions as those reported.

literature (Melliou et al., 2005; Masuda et al., 1998). Control samples (buffered aqueous solutions with DMSO and

The purities of all compounds were evaluated by HPLC prior the without any test compound) were used for all of the plant species

bioassay and they all had a purity higher than 98%. assayed.

Evaluated parameters (germination rate, root length and shoot

3.3. Coleoptiles bioassay length) were recorded using a Fitomed system (Castellano et al.,

2001), which allowed automatic data acquisition and statistical

Wheat seeds (Triticum aestivum L. cv. Duro) were sown in 15 cm analysis using its associated software. Data were analyzed

diameter Petri dishes moistened with water and grown in the dark statistically using Welch’s test, with signiﬁcance ﬁxed at 0.01

at 22 1 8C for 3 days (Hancock et al., 1964). The roots and caryopses and 0.05. Results are presented as percentage differences from the

were removed from the shoots. The latter were placed in a Van der control. Zero represents control, positive values represent stimu-

Weij guillotine, and the apical 2 mm were cut off and discarded. The lation, and negative values represent inhibition.

next 4 mm of the coleoptiles were removed and used for the

bioassays. All manipulations were performed under a green safelight Acknowledgements

(Nitsch and Nitsch, 1956). Compounds were predissolved in DMSO

and diluted to the final bioassay concentration with a maximum of This research was supported by the Consejerıá de Economıá,

0.1% DMSO. Parallel controls were also run. The compounds to be Innovacio´ n, Ciencia y Empleo, Junta de Andalucia, Spain (Project

assayed for biological activity were added to test tubes. Phosphate- No. P10-AGR-5822). One of the authors (L. Nebo) thanks CAPES

citrate buffer (2 mL) containing 2% sucrose (Nitsch and Nitsch, 1956) (Coordenaçaõ de Aperfeiçoamento de Pessoal de Nı´vel Superior,

at pH 5.6 was added to each test tube. Five coleoptiles were placed in Brazil) and FAPESP (Fundac¸a˜o de Amparo a` Pesquisa do Estado de

each test tube (three tubes per dilution) and the tubes were rotated at Sa˜o Paulo, Brazil) for scholarships.

0.25 rpm in a roller tube apparatus for 24 h at 22 8C in the dark. The

coleoptiles were measured by digitalization of their images. Data

were statistically analyzed using Welch’s test (Martı´n Andre´s and References

Luna del Castillo, 1990). Data are presented as percentage differences

Al-Amiery, A.A., Kadhum, A.A.H., Mohamad, A.B., 2012. Antifungal activities of new

from control. Thus, zero represents the control, positive values

coumarins. Molecules 17, 5713–5723.

represent stimulation of the studied parameter, and negative values Anaya, A.L., Macıás-Rubalcava, M., Cruz-Ortega, R., Garcıá-Santana, C., Sańchez-

represent inhibition. Monterrubio, P.N., Herna´ndez-Bautista, B.E., Mata, R., 2005. Allelochemicals

from Stauranthus perforatus, a Rutaceous tree of the Yucatan Peninsula, Mexico.

Phytochemistry 66, 487–494.

3.4. Phytotoxicity bioassay

Anand, P., Singh, B., Singh, N., 2012. A review on coumarins as acetylcholinesterase

inhibitors for Alzheimer’s disease. Bioorg. Med. Chem. 20, 1175–1180.

Arruda, A.C., Vieira, P.C., Fernandes, J.B., Silva, M.F.G.F., 1993. Further pyrano

The selection of target plants was based on an optimization

ﬂavones from Neoraputia alba. J. Braz. Chem. Soc. 4, 80–83.

process developed by us in our search for a standard phytotoxicity

Benavente-Garcı´a, O., Castillo, J., 2008. Update on uses and properties of Citrus

bioassay (Macıás et al., 2000). Several Standard Target Species Flavonoids: new findings in anticancer, cardiovascular, and anti-inflammatory

activity. J. Agric. Food Chem. 56, 6185–6205.

(STS) were proposed, including monocots T. aestivum L. (wheat)

Bermu´ dez-Torres, K., Herrera, J.M., Brito, R.F., Wink, M., Legal, L., 2009. Activity of

and A. cepa L. (onion) and dicots L. esculentum Will. (tomato), L.

quinolizidine alkaloids from three Mexican Lupinus against the lepidopteran

sativum L. (cress) and L. sativa L. (lettuce), which were assayed for crop pest Spodoptera frugiperda. BioControl 54, 459–466.

Braga, P.A.C., Severino, V.G.P., Freitas, S.D.L., Silva, M.F.G.F., Fernandes, J.B., Vieira,

this study. Bioassays were conducted using Petri dishes (50 mm

P.C., Pirani, J.R., Groppo, M., 2012. Dihydrocinnamic acid derivatives from Hortia

diameter), with one sheet of Whatman No. 1 ﬁlter paper as

species and their chemotaxonomic value in the Rutaceae. Biochem. Syst. Ecol.

support. Germination and growth were conducted in aqueous 43, 142–151.

Castellano, D., Macı´as, F. A., Castellano, M., Cambronero, R., 2001. Spanish Patent N8 solutions at controlled pH using 10 M 2-[N-morpholino]etha-

P9901565.

nesulfonic acid (MES) and 1 M NaOH (pH 6.0).

Cazal, C.M., Domingues, V.C., Batalha˜o, J.R., Bueno, O.C., Rodrigues-Filho, E., Silva,

Compounds to be assayed were dissolved in DMSO (0.1, 0.02, M.F.G.F., Vieira, P.C., Fernandes, J.B., 2009. Isolation of xanthyletin, an inhibitor

0.01 and 0.002 M) and these solutions were diluted with buffer of ants’ symbiotic fungus, by high-speed couter-current chromatography. J.

Chromatogr. A 1216, 4307–4312.

(5 ml DMSO solution/ml buffer) so that test concentrations for each

4 4 5 5 Champagne, D.E., Koul, O., Isman, M.B., Scudder, G.G.E., Towers, G.H.N., 1992.

compound (3 10 , 10 , 3 10 and 10 M) were achieved.

Biological activity of limonoids from the Rutales. Phytochemistry 31, 377–394.

This procedure facilitated the solubility of the assayed compounds. Cushnie, T.P.T., Lamb, A.J., 2011. Recent advances in understanding the antibacterial

properties of ﬂavonoids. Int. J. Antimicrob. Ag. 38, 99–107.

The number of seeds in each Petri dish depended on the seed size.

Cutler, H.G., 1984. Fresh look at the wheat coleoptile bioassay. In: Proceedings of

20 seeds were used for tomato, lettuce, cress and onion. Treatment, th

the 11 Annual Meeting of the Plant Growth Regulator Society of America.

control or internal reference solution (1 ml) was added to each PGRSA, Boston, MS, USA, pp. 1–9.

232 L. Nebo et al. / Phytochemistry Letters 8 (2014) 226–232

Cutler, S.J., Hoagland, R.E., Cutler, H.G., 2000. Evaluation of selected pharmaceuticals Michael, J.P., 2005. Quinoline, quinazoline and acridone alkaloids. Nat. Prod. Rep. 22,

as potential herbicides: Bridging the gap between agrochemicals and pharma- 627–646.

ceuticals. In: Narwal, s.s., Hoagland, R.E., Dilday, R.H., Reigosa Roger, M.J. (Eds.), Mishra, B.B., Kale, R.R., Singh, R.K., Tiwan, V.K., 2009. Alkaloids: future prospective to

Allelopathy in Ecological Agriculture and Forestry. Springer, Dordrecht, The combat leishmaniasis. Fitoterapia 80, 81–90.

Netherlands, pp. 129–137. Moraes, V.R.S., Tomazela, D.M., Ferracin, R.J., Garcia, C.F., Sannomiya, M., Soriano,

Da Silva, M.F.G.F., Gottlieb, O.R., Dreyer, D.L., 1984. Evolution of Limonoids in the M.P.C., Silva, M.F.G.F., Vieira, P.C., Fernandes, J.B., Rodrigues Filho, E., Magalha˜es,

Meliaceae. Biochem. Syst. Ecol. 12, 299–310. E.G., Magalha˜es, A.F., Pimenta, E.F., Souza, D.H.F., Glaucius, O., 2003. Enzymatic

Da Silva, M.F.G.F., Gottlieb, O.R., 1987. Evolution of quassinoids and limonoids in the inhibition studies of selected ﬂavonoids and chemosystematic signiﬁcance of

Rutales. Biochem. Syst. Ecol. 15, 85–103. polymethoxylated ﬂavonoids and quinolone alkaloids in Neoraputia (Rutaceae).

Dayan, F.E., Cantrell, C.L., Duke, S.O., 2009. Natural products in crop protection. J. Braz. Chem. 14, 380–387.

Bioorg. Med. Chem. 17, 4022–4034. Mota, K.S.L., Dias, G.E.N., Pinto, M.E.F., Luiz-Ferreira, A., Souza-Brito, A.R.M., Hiruma-

Duarte, R.A., Mello, E.R., Araki, C., Bolzani, V.S., Silva, D.H.S., Regasini, L.O., Silva, Lima, C.A., Barbosa-Filho, J.M., Batista, L.M., 2009. Flavonoids with gastropro-

T.G.A., Morais, M.C.C., Ximenes, V.F., Soares, C.P., 2010. Alkaloids extracted from tective activity. Molecules 14, 979–1012.

Pterogyne nitens induce apoptosis in malignant breast cell line. Tumor Biol. 31, Ngadjui, B.T., Dongo, E., Happi, E.N., Bezabih, M., Abegaz, B.M., 1998. Prenylated

513–522. ﬂavones and phenylpropanoid derivatives from roots of Dorstenia psilurus.

Ferracin, R.J., Silva, M.F.G.F., Fernandes, J.B., Vieira, P.C., 1998. Flavonoids from the Phytochemistry 48, 733–737.

fruits of Murraya paniculata. Phytochemistry 47, 393–396. Nitsch, J.P., Nitsch, C., 1956. Studies on the growth of coleoptile and ﬁrst internode

Garcı´a-Lafuente, A., Guillamo´ n, E., Villares, A., Rostagno, M.A., Martı´nez, J.A., 2009. sections. A new sensitive, straight-growth test for auxins. Plant. Physiol. 31,

Flavonoids as anti-inﬂammatory agents: implications in cancer and cardiovas- 94–111.

cular disease. Inﬂamm. Res. 58, 537–552. Oliveira, F.M., Santana, A.E.G., Conserva, L.M., Maia, J.G.S., Guilhon, G.M.P., 1996.

Godoy, M.F.P., Victor, S.R., Bellini, A.M., Guerreiro, G., Rocha, W.C., Bueno, O.C., Alkaloids and coumarins from Esenbeckia species. Phytochemistry 41, 647–649.

Hebling, M.J.A., Bacci Jr., M., Silva, M.F.G.F., Vieira, P.C., Fernandes, J.B., Pagnocca, Passador, E.A.P., Silva, M.F.G.F., Rodrigues Filho, E., Fernandes, J.B., Vieira, P.C., Pirani,

F.C., 2005. Inhibition of the symbiotic fungus of leaf-cutting ants by coumarins. J.R., 1997. A pyrano chalcone and a ﬂavanone from Neoraputia magnı´ﬁca.

J. Brazil Chem. Soc. 16, 669–672. Phytochemistry 45, 1533–1537.

Guan, A., Liu, C., Li, M., Zhang, H., Li, A., Li, Z., 2011. Design, synthesis and structure- Paulini, H., Waibel, R., Schimmer, O., 1989. Mutagenicity and structure-mutagenic-

activity relationship of novel coumarin derivatives. Pest Manag. Sci. 67, ity relationships of furoquinolines, naturally occurring alkaloids of the Ruta-

647–655. ceae. Mut. Res. 277, 179–182.

Hale, A.L., Meepagala, K.M., Oliva, A., Aliotta, G., Duke, S.O., 2004. Phytotoxins from Pavela, R., Vrchotova´, N., 2013. Insecticidal effect of furanocoumarins from fruits of

the leaves of Ruta graveolens. J. Agric. Food Chem. 52, 3345–3349. Angelica archangelic L. against larvae Spodoptera littoralis Boisd. Ind. Crop Prod.

Hancock, C.R., Barlow, H.W., Lacey, H.J., 1964. The east malling coleoptile straight 43, 33–39.

growth test method. J. Exp. Bot. 15, 166–176. Pick, A., Muller, H., Mayer, R., Haenisch, B., Pajeva, I.K., Weight, M., Bo¨nisch, H.,

Harada, K., Kubo, H., Tomigahara, Y., Nishioka, K., Takahashi, J., Momose, M., Inoue, Mu¨ ller, C.E., Wiese, M., 2011. Structure–activity relationships of ﬂavonoids as

S., Kojima, A., 2010. Coumarins as novel 17-b-hydroxysteroid dehydrogenase inhibitors of breast cancer resistance protein (BCRP). Bioorg. Med. Chem. 19,

type 3 inhibitors for potential treatment of prostate cancer. Bioorg. Med. Chem. 2090–2102.

Lett. 20, 272–275. Salas, M.P., Ce´liz, G., Geronazzo, H., Daz, M., Resnik, S.L., 2011. Antifungal activity of

Hongwei, Y., Bogang, L., Xiaozhen, C., Changsong, L., Guolin, Z., 2010. Chemical study natural and enzymatically-modiﬁed ﬂavonoids isolated from citrus species.

on Evodia vestita. Chin. J. Appl. Environ. Biol. 16, 72–75. Food Chem. 124, 1411–1415.

Jacyno, J.M., Cutler, H.G., 1993. Detection of herbicidal properties: scope, and Sampaio, O.M., Silva, M.F.G.F., Veiga, T.A.M., King-Dı´az, B., Lotina-Hennsen, B., 2012.

limitations of the etiolated wheat coleoptiles bioassay. PGRSA Quarterly 21, Avaliaçaõ de furanocumarinas como inibidores da fotossıńtese atrave´s de

15–24. ensaios de ﬂuoresceˆncia da Cloroﬁla a. Quı´m. Nova 11, 2115–2118.

Joosten, L., Veen, J.A., 2011. Defensive properties of pyrrolizidine alkaloids against Santos, D.A.P., 2005. Busca de metabo´ litos bioativos em plantas das famı´lias

microorganisms. Phytochem. Rev. 10, 127–136. Bignoniaceae e Rutaceae contra parasitas causadores de doenc¸as tropicais.

Kim, H.P., Son, K.H., Chang, H.W., Kang, S.S., 2004. Anti-inflammatory plant flavo- PhD Thesis, Universidade Federal de Saõ Carlos, Maio.

noids and cellular action mechanisms. J. Pharmacol. Sci. 96, 229–245. Santos, D.A.P., Vieira, P.C., Silva, M.F.G.F., Fernandes, J.B., Rattray, L., Croft, S.I., 2009.

Kumar, V., Vallipuram, K., Adebajo, A.C., Reisch, J., 1995. 2,7-Dihydroxy-3-formyl-1- Antiparasitic activities of acridone alkaloids from Swinglea glutinosa (BI.). Merr.

0 0

(3 -methyl-2 -butenyl)carbazole from Clausena lansium. Phytochemistry 40, J. Braz. Chem. Soc. 20, 644–651.

1563–1565. Sardari, S., Mori, Y., Horita, K., Micetich, R.G., Nishibe, S., Daneshtalab, M., 1999.

Leite, A.C., Ambrozin, A.R.P., Castilho, M.S., Vieira, P.C., Fernandes, J.B., Oliva, G., Synthesis and antifungal activity of coumarins and angular furanocoumarins.

Silva, M.F.G.F., Thiemann, O.H., Lima, M.I.S., Pirani, J.R., 2009. Screening of Bioorg. Med. Chem. 7, 1933–1940.

Trypanosoma cruzi glycosomal glyceraldehyde-3-phosphate dehydrogenase Silva, L.F.R., Montoia, A., Amorim, R.C.N., Melo, M.R., Henrique, M.C., Numomura,

enzyme inhibitors. Braz. J. Pharmacogn. 19, 1–6. S.M., Costa, M.R.F., Andrade Neto, V.F., Costa, D.S., Dantas, G., Lavrado, J.,

Li, Z., Gu, Y., Irwin, D., Sheridan, J., Clough, J., Chen, P., Peng, S., Yang, Y., Guo, Y., 2009. Moreira, R., Paulo, A., Pinto, A.C., Tadei, W.P., Zacardi, R.S., Eberlin, M.N., Pohlit,

Further Daphniphyllum alkaloids with insecticidal activity from the bark of A.M., 2012. Comparative in vitro and in vivo antimalarial activity of the indole

Daphniphyllum macropodum MiQ. Chem. Biodiv. 6, 1744–1750. alkaloids ellipticine, olivacine, cryptolepine and a synthetic cryptolepine ana-

Macı´as, F.A., Galindo, J.C.G., Massanet, G.M., Rodriguez-Luis, F., Zubia, E., 1993. log. Phytomedicine 20, 71–76.

Allelochemicals from Pilocarpus goudotianus leaves. J. Chem. Ecol. 19, 1371– Souto, A.L., Tavares, J.F., Silva, M.S., Diniz, M.F.F.M., Athayde-Filho, P.F., Barbosa-

1379. Filho, J.M., 2011. Anti-inﬂammatory activity of alkaloids: An update from 2000

Macı´as, F.A., Molinillo, J.M.G., Torres, A., Varela, R.M., Castellano, D., 1997. Bioactive to 2010. Molecules 16, 8515–8534.

ﬂavonoids from Helianthus annuus cultivars. Phytochemistry 45, 683–687. Souza, S.M., Delle-Monache, F., Smaˆnia Jr., A., 2005. Antibacterial activity of cou-

Macı´as, F.A., Castellano, D., Molinillo, J.M.G., 2000. Search for a standard phytotoxic marins. Z. Naturforsch. 60, 693–700.

bioassay for allelochemicals. Selection of standard target species. J. Agric. Food Tan, Q., Luo, X., 2011. Meliaceous limonoids: chemistry and biological activities.

Chem. 48, 2512–2521. Chem. Rev. 111, 7437–7522.

Macı´as, F.A., Molinillo, J.M.G., Varela, R.M., Galindo, J.C.G., 2007. Allelopathy – a Tomazela, D.M., Pupo, M.T., Passador, E.A.P., Silva, M.F.G.F., Vieira, P.C., Fernandes,

natural alternative for weed control. Pest Manag. Sci. 63, 327–348. J.B., Rodrigues Filho, E., Oliva, G., Pirani, J.R., 2000. Pyrano chalcones and a

Marchand, L.L., 2002. Cancer preventive effects of flavonoids – a review. Biomed. flavone from Neoraputia magnifica and their Trypanosoma cruzi glycosomal

Pharmacother. 56, 296–301. glyceraldehyde-3-phosphate dehydrogenase-inhibitory activities. Phytochem-

Martı´n Andre´s, A., Luna Del Castillo, J.D., 1990. Bioestadı´stica para las Ciencias de la istry 55, 643–651.

Salud, 3rd ed. Norma, Madrid. Treutter, D., 2006. Signiﬁcance of ﬂavonoids in plant resistance: a review. Environ.

Masuda, T., Takasugi, M., Aneta, M., 1998. Psoralen and other linear furanocoumar- Chem. Lett. 4, 147–157.

ins as phytoalexins in Glehnia littoralis. Phytochemistry 47, 13–16. Velozo, E.S., Oliveira, D.J., Arruda, A.C., Vieira, P.C., Fernandes, J.B., Silva, M.F.G.F.,

Mbeunkui, F., Grace, M.H., Lategan, C., Smith, P.J., Raskin, I., Lila, M.A., 2012. In vitro Caracelli, I., Zukerman-Schpector, J., 1997. Rauianin, a new coumarin from Rauia

antiplasmodial activity of indole alkaloids from the stem bark of Geissospermum resinosa. Nat. Prod. Res. 9, 237–244.

vellosii. J. Ethnopharmacol. 139, 471–477. Wolfe, K.L., Liu, R.H., 2008. Structure-activity relationships of ﬂavonoids in the

Melliou, E., Magiatis, P., Mitaku, S., Skaltsounis, A., Chinou, E., Chinou, I., 2005. cellular antioxidant activity assay. J. Agric. Food Chem. 56, 8404–8411.

Natural and synthetic 2,2-dimethylpyranocoumarins with antibacterial activi- Zhai, H., Zhao, C., Zhang, N., Jin, M., Tang, S., Qin, N., Kong, D., Duan, H., 2012.

ty. J. Nat. Prod. 68, 78–82. Alkaloids from Pachysandra terminalis inhibit breast cancer invasion and have

Michael, J.P., 2007. Quinoline, quinazoline and acridone alkaloids. Nat. Prod. Rep. 24, potential for development as antimetastasis therapeutic agents. J. Nat. Prod. 75, 223–246. 1305–1311.